Th17 cells constitute a population of T helper cells that secrete the pro-inflammatory cytokine IL-17. These cells have been proposed to be key in the development of a range of human diseases including various autoimmune conditions, inflammation, allergy and development of the tumor microenvironment (see Park et al., A distinct lineage of CD4 T cells regulate tissue inflammation by producing IL-17, Nat. Immunol. 6:1133-1141, 2005; Qi et al., Correlation between Th17 cells and tumor microenvironment, Cell. Immunol. 285: 18-22, 2013).
Besides the preclinical studies, there are clinical data correlating enhanced IL-17 levels and increased frequencies of Th17 in a variety of human diseases. Elevation of IL-17 has been detected in the sera and biopsies of rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) patients (see Agarwal et al., Interleukin 17 levels are increased in juvenile idiopathic arthritis synovial fluid and induce synovial fibroblasts to produce proinflammatory cytokines and matrix metalloproteinases, J. Rhematology, 35:515-519, 2008; Wong et al., Hyperproduction of IL-23 and IL-17 in patients with systemic lupus erythematosus: implications for Th17-mediated inflammation in auto-immunity, Clin. Immunol. 127:385-393, 2008). IL-17A promotes ex vivo induction of proinflammatory cytokines by synoviocytes from RA patients (see Chabaud et. al., Enhancing effect of IL-17 on IL-1-induced IL-6 and leukemia inhibitory factor production by rheumatoid arthritis synoviocytes and its regulation by Th2 cytokines, J. Immunol. 161:409-414, 1998.) and a humanized anti-IL-17 antibody has been used for a proof-of-concept clinical trial for RA patients with positive results (see Genovese et al., LY2439821, a humanized anti-interleukin-17 monoclonal antibody, in the treatment of patients with rheumatoid arthritis: a phase I randomized, double-blind, placebo-con-trolled, proof-of-concept study, Arthr. & Rheum. 64: 929-979, 2010). In addition, RA patients treated with humanized anti-IL17 antibody showed reductions in symptoms similar to those produced by the approved anti-TNF antibody infliximab thus providing additional evidence that IL-17 is an important mediator of inflammatory autoimmune diseases.
In multiple sclerosis (MS) patients, the level of messenger RNA for IL-17 in the monocytes from cerebrospinal fluid is elevated and myelin reactive Th17 cells are enriched (see Venken et al., Memory CD4+CD127 high T cells from patients with multiple sclerosis produce IL-17 in response to myelin antigens, J. Neuroimmunol. 226: 185-191, 2010). In psoriatic skin, IL-17A, IL-22 and IL-23 are all elevated and IL-17A induces the expression of genes involved in psoriasis by human keratinocytes (see Res et al., Overrepresentation of IL-17A and IL-22 producing CD8 T cells in lesional skin suggests their involvement in the pathogenesis of psoriasis, PLOS 5: Article ID e14108, 2010; Wilson et al., Development, cytokine profile and function of human interleukin 17-producing helper T cells, Nat. Immunol. 8: 950-957, 2007). An elevation of IL-17 positive cells has also been detected in the affected areas of the gut of Crohn's disease patients compared with unaffected areas (see Annunziato et al., Phenotypic and functional features of human Th17 cells, J. Exp. Med. 204:1849-18-61, 2007). Taken together these human data support modulation of Th17 cells as a therapeutic intervention in a variety of diseases including MS, RA, SLE, psoriasis, and Crohn's disease.
Kinases play critical roles in signaling pathways controlling fundamental cellular processes such as proliferation, differentiation, and death (apoptosis). Two kinases, in particular, ITK and RLK (also referred to as TXK) have been shown to be important for T cell function. In ITK −/− (ITK deleted) CD4+ cells differentiated in vitro into Th17 cells the levels of intracellular IL-17A levels were greatly reduced. It has been observed that differentiation of Th17 cells is impacted as well (see Gomez-Rodriguez et al., Differential expression of IL-17A and IL-17F is coupled to TCR signaling via ITK-mediated regulation of NFATc1, Immunity 31:587-597, 2009). In addition to the decreased differentiation and activation of Th17 cells in the absence of ITK, T cells from these ITK−/− mice show enhanced differentiation of Treg cells that modulate inflammation and prevent development of autoimmunity (see Gomez-Rodriguez et al., Itk-mediated integration of T cell receptor and cytokine signaling regulates the balance between Th17 and regulatory T cells, J. Exp. Med. 10.1084/jem.20131459. 2014). Thus in the absence of ITK, there appears to be a shift in the balance of Th17 and Treg cells that would favor suppression of autoimmune disease. Indeed, when T cells from ITK−/− mice are transferred into a model of T cell mediated colitis, the enhanced Treg capacity suppresses development of the colitits disease model. In yet another in vivo model of IL-17A associated inflammation, antigen-stimulated allergic asthma, the levels of IL-17A in the lungs of ITK−/− mice were reduced in the absence of ITK as well. It is noteworthy that this is not a developmental defect as is sometimes the case for non-induced, germline gene knockout models because the defect in IL-17A expression can be rescued by re-expression of ITK in pre-activated cells suggesting that modulation of ITK with a small molecule inhibitor could affect Th17 differentiation and therefore the inflammation that results from elevated IL-17 levels.
When the deletion of ITK is combined with deletion of RLK in double knockout mice, the effects on TCR signaling and IL-17A production are more severe than in the single knockout mice. Additionally, in the absence of both ITK and RLK, T-cell receptor signaling is impaired, with defects in mitogen-activated protein kinase activation, Ca2+ mobilization, and actin polymerization (see Berg et al., Tec family kinases in T lymphocyte development and function, J. Exp. Med, 23:549-600, 2005).
In general, the effects of knocking out both ITK and RLK in mice causes more profound changes in TCR mediated signaling, T cell functions and production of IL17A than deletion of either kinase alone (see Schaeffer et al., Requirement for Tec kinases RLK and ITK in T cell receptor signaling and immunity, Science, 284:638-641, 1999; Gomez-Rodriguez et al., Differential expression of IL-17A and IL-17F is coupled to TCR signaling via ITK-mediated regulation of NFATc1, Immunit, 31:587-597, 2009; Felices and Berg, The Tec kinases ITK and RLK regulate NKT cell maturation, cytokine production and survival, J. Immunol., 180:3007-3018, 2008). Therefore a compound that is a potent inhibitor of both ITK and RLK kinases could be expected to have different effects and enhanced efficacy on modulation of TCR-induced signaling, including effects on IL-17 mechanisms than one that blocks only one of RLK and ITK kinase.